This project will study the metabolic function and regulation of mitochondrial CD36, a fatty acid binding protein that has been shown to facilitate transport of long-chain fatty-acids (LCFA) across cellular membranes. Although the role of CD36 in regulating fatty acid transport across the plasma membrane has been previously studied, only recently was it demonstrated that CD36 also participates in fatty acid uptake into muscle mitochondria. The physiological and pathophysiological relevance of this observation is still poorly understood. Using recombinant adenovirus technology we will examine the impact of CD36 over- expression and silencing on mitochondrial fuel metabolism and insulin sensitivity in rat L6 myocytes. Changes in mitochondrial metabolism will be assessed by classic functional assays as well as mass spectrometry-based metabolic profiling. We hypothesis that increased CD36 will promote uncontrolled beta- oxidation, mitochondrial stress and increased susceptibility of cultured myocytes to fatty-acid induced insulin resistance. Using whole animal models, we will also test our hypothesis that obesity and insulin resistant states promote mislocalization of CD36 to the mitochondrial membrane. Finally, we will characterize changes in the intracellular itinerary of CD36 in response to insulin and contraction, both recently show to modulate the function of CD36 by altering its localization. Results from these studies will greatly enhance our understanding of the mechanisms of CD36 regulation, as well as the metabolic consequences of its mis- regulation. Relevance: The epidemics of obesity and diabetes are closely associated with lipid dysregulation, both systemically and within peripheral tissues such as skeletal muscle. CD36, a mitochondrial-localized fatty-acid transport protein, has been shown to be up-regulated in diabetic states and has been implicated as a candidate mediator of the pathology. The proposed research will increase our understanding of the specific role of CD36 in regulating mitochondrial function and dysfunction as well as its potential connection to the development of type 2 diabetes. [unreadable] [unreadable] [unreadable]